Head and neck cancer develops over many years and mechanisms that control and regulate progression are poorly defined. During this time period, progressing cells are surrounded by normal cells and communicate with subjacent stroma within 3-dimensional tissue structure. It is necessary to consider both the spatial organization of cells and their interactions with their environment, defined as microenvironment, to develop models for early stages of cancer (premalignancy). We hypothesize that in early stages of promotion and progression, the microenvironment regulates the interactions between premalignant oral epithelial (POE) cells and adjacent normal oral epithelial (NOE) cells, leading to expansion of POE at the expense of NOE. Secondly, we hypothesize that the mechanisms involved in POE expansion include regulation by the microenvironment of the basic homeostatic mechanisms (proliferation, differentiation and apoptosis) that control and maintain tissue structure and regulate cell behavior. Primary cultures of NOE, defined as normal through comparative genomic hybridization, and unique POE cell lines developed from dysplastic leukoplakia, will be used to model NOE:POE interactions. Microenvironment will be manipulated by varying extracellular matrix material (ECM) and fibroblasts, as mediators of mesenchymal information. Three coculture systems of increasing complexity will be used to model early lesions. 1) NOE will be confronted with POE on coverslips and microenvironmental complexity increased by addition of various ECMs and fibroblasts (derived from normal gingiva, dysplastic leukoplakia or tumor). Videomicroscopy will provide real time information on how the microenvironment affects the 'social' behavior of NOE:POE cells and their proliferative, differentiative and apoptotic response. Immunostaining procedures will provide quantitative values for these homeostatic processes. 2) Organotypic cultures (OC) with 3- dimensional histoarchitechture, possessing a normal-dysplastic junction, will be used to define mechanisms through which NOE:POE interactions affect the homeostatic mechanisms controlling tissue structure. 3) In vivo grafting of NOE/POE collagen gels onto nude mice will generate an in vivo normal-dysplastic tissue and extend the time frame for defining how NOE:POE interactions control tissue structure. Manipulation of integrins with adenoviral infections will further define mechanisms regulating tissue structure since integrins are fundamental to epithelial biology. Understanding the role of the microenvironment in models for premalignancy may lead to new therapeutic and preventive strategies for treating and reversing early disease.